Abstract

Pro-carcinogens, such as benzoapyrene (BaP), that are exogenous ligands of the aromatic hydrocarbon receptor may influence the susceptibility of target-cell populations through the up-regulation of cytochrome P450 (CYP) mixed function oxidases. We examined whether the growth kinetics of MCF-7 cells might determine the level of up-regulation of CYP1A1, CYP1A2 or CYP1B1 by BaP, and whether this could then influence subsequent levels of DNA damage. Cell cultures manipulated to be G0/G1-phase concentrated, S-phase concentrated or G2/M-phase concentrated were treated with BaP and the expression levels of CYP1A1, CYP1A2, CYP1B1, cyclin-dependent kinase inhibitor 1A CDKN1A (P21WAF1/CIP1), B-cell leukaemia/lymphoma-2 (BCL-2), and Bcl-2-associated X levels were determined. Levels of DNA damage were measured as DNA single-strand breaks (SSBs) by the alkaline single-cell gel electrophoresis (comet) assay or as DNA adducts by 32P-postlabelling analysis. BaP-induced up-regulation of CYP1A1 was {\ensuremath{>}}100-fold in S-phase-concentrated cells, but in G0/G1-phase- or G2/M-phase-concentrated cultures up-regulation occurred to a significantly lower extent. Consistent with this, BaP-treated S-phase-concentrated cultures exhibited markedly up-regulated P21WAF1/CIP1, higher levels of dose-related increases in DNA SSBs, and increased DNA adduct levels presumably as a result of CYP1A1-mediated activation of BaP to BaP-diol-epoxide compared with the cultures enriched for the other cell cycle phases. Growth kinetics in vitro may be an important predeterminant of susceptibility to an exogenous pro-carcinogen in short-term test systems and these findings have important implications when extrapolating such results to a particular target-cell population in vivo.